Carotenoid and chlorophyll content of Vitis vinifera cv. Merlot grapes during ripening with reference to variability in grapevine water status and vigour

ENGLISH ABSTRACT: Previous research has shown that carotenoids are precursors of C13-norisoprenoid aroma
compounds in wine. C13-norisoprenoids have low threshold values in wine with the most
prominent C13-norisoprenoids being β-damascanone and β-ionone which contribute honey and
floral like aroma to wine. Chlorophyll and its derivates have also been detected in wine with
potential to be precursors to aroma compounds.
Apart from the contribution of these pigments to wine aroma and quality they are vital role
players in photosynthesis and are widely found in plants and plant products. The main functions
of these pigments in plants are light collection and light-protection.
Research has shown that environmental conditions, climate, light exposure of bunches and
soil water deficit influence the carotenoid content of grape berries. Furthermore the
concentration of carotenoids and chlorophylls has also been shown to differ between cultivars.
No research in this regard has been done on Merlot grape berries.
With this in mind, the aim of this study was to evaluate the effect of vigour and soil water
content on the evolution of carotenoids and chlorophylls through ripening of grape berries from
the cv. Merlot. However, when looking at methods to analyse carotenoids and chlorophylls in
berry tissue, especially lyophilised tissue, there were no readily available methods. Thus, an
extraction method to identify and quantify the carotenoid and chlorophyll profile of lyophilised
tissue from unripe (green) to ripe (red) Merlot grape berries was needed. In this study the RPHPLC
method of Taylor et al. (2006) for carotenoids and the extraction method of Mendes-Pinto
et al. (2004) were adapted to analyse both carotenoids and chlorophylls in lyophilised grape
tissue. The RP-HPLC method baseline separated all the carotenoids and chlorophylls and their
derivatives. Recovery of standards from mock extractions was high, indicating that the
extraction procedure was acceptable. However, extraction recovery tested in the matrix of the
grape tissue showed less promising results due to the high acid content of grape tissue.
Violaxanthin, neoxanthin and the chlorophylls were especially sensitive to low pH conditions
which facilitated their degradation. The degradation products of these compounds under acidic
conditions were identified as pheophytin a, b, chlorophillide a, pyropheophytin b, cisviolaxanthin,
cis-neoxanthin, neochrome, mutatoxanthin and luteoxanthin. There is a possibility
that some degradation products were already present in the tissue due to lyophilisation (since
the water in the berry was then removed and the acid concentrated). More work is needed to
investigate the effect of lyophilisation and storage on the composition of grape tissue of different
maturity. The extraction method for grape berry tissue at different ripening stages should also
be optimised further to effectively neutralise tissue acidity, without compromising the extraction
of carotenoids significantly, in especially green berry tissue. The question as to whether cisisomers
and chlorophyll degradation products are naturally present in grape berries or are
formed during sampling and processing remains unanswered in the current study. This study confirmed that in general carotenoids and chlorophylls decrease on a per berry
(μg/berry) and concentration (μg/g) basis from veraison to harvest. Furthermore, this study was
inconclusive in showing that vigour differences have an effect on the rate of
synthesis/degradation of carotenoids, chlorophyll and some other ripening parameters, namely
malic acid, total glucose and fructose, total tannin and total anthocyanin, from pre-veraison (pea
size) to harvest. Additionally, no significant effect of soil water content on carotenoids,
chlorophylls and ripeness parameters was found in this study, most likely due the fact that high
soil water capacity was found in lower soil layers which may have prevented significant
differences in grapevine water status. Experimental plots selected for vigour differences based
on normalised difference vegetation index (NDVI) images, pruning mass and soil water
measurements by means of a neutron probe, showed significant differences in soil water
content in only the first 30 cm of the soil for the ripening seasons studied. Predawn plant water
potential measurements, however, indicated that none of the experimental vines experienced
severe water stress which was previously shown to effect carotenoid content of grapes.
The carotenoid 5,8-epoxy--carotene was quantified for the first time in grapes and
represents a significant amount of the total carotenoids present at harvest. All the carotenoids
and chlorophylls except -carotene appeared to be sensitive to seasonal variation in climatic
conditions. Lutein and β-carotene were found to be the most abundant carotenoids present in
Merlot grape berries together with chlorophyll a for both seasons studied. The values of these
carotenoids also correlated well with previous research. However, chlorophyll a was found in
much larger quantities in Merlot berries compared to reported data. This is possibly because in
this study the chlorophyll degradation products were included in the calculation of chlorophyll a.
Multivariate analysis showed promising preliminary prediction models (with correlation
values of above 0.8 for both seasons analysed) for the prediction of the concentration of
ripeness parameters (glucose, fructose, malic acid, total tannins and anthocyanins) with
carotenoid and chlorophyll content. This result highlights the opportunity for the development of
a rapid non-destructive method to measure carotenoids and chlorophylls in berries which in turn
can predict optimal ripeness. Furthermore, since carotenoids are the precursors to C13-
norisoprenoid aroma compounds in wine a preview of the potential contribution of these aromas
to wine might be evaluated. Further research is necessary to investigate the possibility of
building and validating such models.